Early detection of cancer of the mouth and throat (head and neck squamous cell carcinoma [HNSCC]) would result in substantial improvement in survival for the 40,000 Americans diagnosed with this disease each year while reducing morbidity associated with treatment, Changes in DNA that underly tumorigenesis are ideal molecular markers for highly sensitive and specific approaches to early detection. We have had promising results in pilot projects for detection of HNSCC in oral rinses (saliva) and serum of affected individuals using hypermethylation markers identified during the first funding period by Project 1. In the proposed new funding period, it is anticipated that the number of promising markers identified in Project 1 will double or triple . In order to produce an accurate, useful and cost-effective detection test, it will be necessary to combine markers that have shown initial promise into detection panels. However, the presence of some markers in saliva and serum of healthy control subjects requires that detection panels be compartment specific.We will assemble and verify candidate markers and panels through the assessment of tumor-specific hypermethylation of promoter regions in tumors, premalignant lesions, exfoliated cells in oral rinse specimens, and serum. Quantitative methylations specific PCR which is amenable to high-throughput application will be employed. The prevelance of alterations in sets of tumors will be assessed, followed by evaluation of the presence of identical markers in clinical specimens (saliva and serum). Simple sensitivity and specificity estimates will be explored in various panel combinations. The proposed panels of markerswill then be tested in samples from subjects enrolled in premalignant and post-treatment surveillance studies (aims 2 and 3). Subjects will be recruited at the Johns Hopkins Medical Institutions and the MD Anderson Cancer Center. The utility of marker panels for detection and prediction of recurrence/progression will be explored.